Calcium, carbonic anhydrase and gastric acid secretion.

Previous data concerning the action of calcium (Ca) on gastric acid secretion (GAS) indicated that calcium ions increase GAS elicited by gastrin released through a vagal mechanism, and also by a direct effect on parietal cells. Our research showed that the stimulating effect of calcium on gastric acid secretion can be antagonized by verapamil administration, which reduces gastric acid secretion . In the present study we followed the effect induced by administration of calcium and Ca-chelating agents (disodium EDTA) on gastric acid secretion and on carbonic anhydrase (CA) activity. We selected two groups of healthy volunteers: Group I (n=21) received a single i.v. dose of CaCl2 (15 mg/kg b.w.), whereas Group II (n=22) received a single i.v. dose of disodium EDTA (5 mg/kg b.w.). We determined blood calcium before and after treatment, gastric acid secretion at 2 hours. erythrocyte CA II activity, and CA IV activity in membrane parietal cells, which were isolated from gastric mucosa obtained by endoscopic biopsy. Assessment of carbonic anhydrase activity was achieved by the stopped-flow method. In Group I calcium administration increased blood calcium, HCl output, CA II and CA IV activity as compared to initial values. In Group II, disodium EDTA reduced blood calcium, HCl output, CA II and CA IV activity as compared to initial values. The results demonstrated that increased blood calcium and GAS values after calcium administration correlated with the increase of erythrocyte CA II and parietal cell CA IV activity, while disodium EDTA induced a reversed process. Our results also show that cytosolic CA II and membrane CA IV values are sensitive to calcium changes and they directly depend on these levels. Our data suggest that intra- and extracellular pH changes induced by carbonic anhydrase might account for the modulation of the physiological and pathological secretory processes in the organism.

Indomethacin activates carbonic anhydrase and antagonizes the effect of the specific carbonic anhydrase inhibitor acetazolamide, by a direct mechanism of action.

OBJECTIVES: In this paper we investigated the effect of indomethacin, acetazolamide and their combination in vitro and in vivo on carbonic anhydrase (CA) isozymes. METHOD: In vitro experiments followed the effect of the two substances at concentrations between 10(-8)-10(-4) M on purified human red cell CA I and II as well as on human gastric mucosa CA IV using dose-response relationships. Kinetic studies were also performed. The effects of single and combined administration of indomethacin and acetazolamide on red cell CA and on gastric acid secretion were studied in vivo. RESULTS: Indomethacin, in vitro and in vivo. induces an increase in erythorcyte CA I and CA II activity. Acetazolamide, a specific inhibitor of CA, reduces the activity of CA I and CA II from red cells. Indomethacin completely antagonizes CA activity, i.e. abolishes the inhibitory effect of acetazolamide on CA. In humans, an increase or decrease in erythrocyte CA II activity is correlated with an increase or decrease in gastric acid secretion. CONCLUSIONS: Our results show that indomethacin, a known cyclooxygenase (COX) inhibitor, is also an activator of CA. Our data also prove that indomethacin is not only an activator of CA but also antagonizes the effect of acetazolamide, a specific inhibitor of this enzyme. In view of the role of CA in acid-base balance as well as the fact that an increase or decrease in its activity is accompanied by an increase or decrease in intra- and extracellular pH, our results suggest that: firstly, CA activation induced by indomethacin might cause changes in COX activity; secondly, PGs are synthetized as a consequence of the changes in COX activity, a hypothesis that requires further study.

Vasoconstrictive drugs increase carbonic anhydrase I in vascular smooth muscle while vasodilating drugs reduce the activity of this isozyme by a direct mechanism of action.

Carbonic anhydrase (CA) is a zinc enzyme that catalyses the reversible hydration reaction of CO2 and plays a major role in the acid-base balance. We have previously shown that certain vasoconstrictive therapeutic agents increase CA I activity whereas vasodilating drugs reduce the activity of this isozyme by a direct mechanism of action. In this paper we studied the effect of other vasoconstrictive and vasodilating agents on CA I activity in order to elucidate the involvement of vascular smooth muscle CA I in vasoconstrictive and vasodilating processes. We studied the in vitro effects of noradrenaline, prostaglandin F2 alpha, thromboxane A2, leukotriene B4, angiotensin II, vasopressin, indomethacin, prazosin, hydralazine, clonidine, reserpine, prostaglandin I2, indapamide, furosemide, amlodipine, verapamil and irbesartan on purified human red blood cell CA I and vascular smooth muscle CA I isolated from rabbits. In vivo, we selected six groups of five rabbits each, which were administered the following substances in acute experiments: orciprenaline (group 1), desmopressin (group 2), verapamil (group 3), irbesartan (group 4), acetazolamide (group 5) and placebo (control group). Vascular smooth muscle CA I activity and systolic blood pressure were determined and compared with those of the control group. In vitro results showed that all the vasoconstrictive agents studied increased purified and human erythrocyte CA I activity as well as vascular smooth muscle CA I, while vasodilating substances reduced the activity of isozyme by a direct mechanism of action. The same results obtained in vivo showed that activation of vascular smooth muscle CA I increased blood pressure while its inhibition reduced blood pressure. The results of this study suggest that pHi changes, induced by activating or inhibiting CA I in vascular smooth muscle, might be responsible for changes in vascular tonus.

Catecholamine-induced vasoconstriction is sensitive to carbonic anhydrase I activation.

We studied the relationship between alpha- and beta-adrenergic agonists and the activity of carbonic anhydrase I and II in erythrocyte, clinical and vessel studies. Kinetic studies were performed. Adrenergic agonists increased erythrocyte carbonic anhydrase as follows: adrenaline by 75%, noradrenaline by 68%, isoprenaline by 55%, and orciprenaline by 62%. The kinetic data indicated a non-competitive mechanism of action. In clinical studies carbonic anhydrase I from erythrocytes increased by 87% after noradrenaline administration, by 71% after orciprenaline and by 82% after isoprenaline. The increase in carbonic anhydrase I paralleled the increase in blood pressure. Similar results were obtained in vessel studies on piglet vascular smooth muscle. We believe that adrenergic agonists may have a dual mechanism of action: the first one consists of a catecholamine action on its receptor with the formation of a stimulus-receptor complex. The second mechanism proposed completes the first one. By this second component of the mechanism, the same stimulus directly acts on the carbonic anhydrase I isozyme (that might be functionally coupled with adrenergic receptors), so that its activation ensures an adequate pH for stimulus-receptor coupling for signal transduction into the cell, resulting in vasoconstriction.

Catecholamine-induced vasoconstriction is sensitive to carbonic anhydrase I activation

We studied the relationship between alpha- and beta-adrenergic agonists and the activity of carbonic anhydrase I and II in erythrocyte, clinical and vessel studies. Kinetic studies were performed. Adrenergic agonists increased erythrocyte carbonic anhydrase as follows: adrenaline by 75 percent, noradrenaline by 68 percent, isoprenaline by 55 percent, and orciprenaline by 62 percent. The kinetic data indicated a non-competitive mechanism of action. In clinical studies carbonic anhydrase I from erythrocytes increased by 87 percent after noradrenaline administration, by 71 percent after orciprenaline and by 82 percent after isoprenaline. The increase in carbonic anhydrase I paralleled the increase in blood pressure. Similar results were obtained in vessel studies on piglet vascular smooth muscle. We believe that adrenergic agonists may have a dual mechanism of action: the first one consists of a catecholamine action on its receptor with the formation of a stimulus-receptor complex. The second mechanism proposed completes the first one. By this second component of the mechanism, the same stimulus directly acts on the carbonic anhydrase I isozyme (that might be functionally coupled with adrenergic receptors), so that its activation ensures an adequate pH for stimulus-receptor coupling for signal transduction into the cell, resulting in vasoconstriction

Cysteamine, the most potent ulcerogenic drug known so far, powerfully activates carbonic anhydrase I, II and IV. In vitro and in vivo studies.

Cysteamine is known as the most efficient substance in producing experimental duodenal ulcers reaching 100% after a single dose administration in rats. It is also described the acid hypersecretion and the duodenal mucosa blood flow decrease after cysteamine administration. The mechanism of action is still unknown. Starting from our recent studies which show that carbonic anhydrase (CA) I is involved in vascular changes and CA II and CA IV are involved in the secretory modifications we followed the effect of cysteamine on these CA isozymes. In vitro, we followed the effect of cysteamine on CA I, CA II and CA IV isolated from the gastric mucosa parietal cells and from kidneys using the Maren technique. In vivo, 2 groups of rats Gr.1 (N = 31) received a single s.c. dose of cysteamine, 500 mg/kg b.w. and Gr.2 (N = 32) - s.c. isotonic saline solution. We determined CA I, II and IV activity from parietal cells and renal CA IV activity. CA activity was determined by the stopped-flow method, using a rapid kinetic apparatus HI-TECH SF 51 MX. The results show that in vitro cysteamine activated the purified CA I and CA II, as well as gastric mucosa parietal cell CA IV by a direct mechanism of action. The renal CA IV was not significantly activated by cysteamine. In vivo cysteamine activated gastric mucosa CA I, CA II and CA IV and did not modify the activity of the same isozyme from the kidneys. In vivo and in vitro CA I activation had confirmed our results, and this fact proved the enzyme's involvement in the vasocontrictive process cysteamine-induced. The powerful activation of gastric CA II and CA IV through the H+ source, could explain the HCl excess produced by cysteamine. The absence of cysteamine activating effect on renal CA IV proved organ specificity. The results suggested the involvement of gastric mucosa CA I, II and IV in the experimental cysteamine ulcerogenesis.

Hypotensive effect of calcium channel blockers is parallel with carbonic anhydrase I inhibition.

In this article we studied in vitro and in vivo the effect of calcium channel blockers (verapamil and amlodipine) on erythrocyte carbonic anhydrase I activity, on carbonic anhydrase I isolated from vascular smooth muscles, and on arterial blood pressure values in human beings and in animals. Our in vitro and in vivo results have shown that verapamil and amlodipine are strong inhibitors of carbonic anhydrase I both in erythrocytes (in human beings) and in vascular smooth muscles (in animals). In human beings calcium channel blockers reduce arterial blood pressure in subjects with hypertension and progressively reduce erythrocyte carbonic anhydrase I activity. We assume that verapamil and amlodipine possess a dual mechanism of action: the first mechanism consists of their action on calcium channels, and the second mechanism, proposed by us, shows that verapamil and amlodipine inhibit vascular smooth muscle carbonic anhydrase I activity with consecutive pH increase. The increase of pH might be an additional factor involved in intracellular calcium influx through calcium channels. This dual mechanism of action would bring new data regarding the hypotensive effect of verapamil and amlodipine, effects that might also be parallel and dependent on carbonic anhydrase I inhibition.

Carbonic anhydrase I inhibition by nitric oxide: implications for mediation of the hypercapnia-induced vasodilator response.

1. At present, CO2 is considered to be the most important factor in regulating cerebral blood flow by modification of the interstitial fluid and extracellular pH, but the mechanism by which hypercapnia produces vasodilation is still controversial. In the present paper we investigated the effect of hypercapnia on carbonic anhydrase (CA) activity. We also studied the combined effects of CO2 with either indomethacin or an L-arginine analogue on CA activity. 2. Nine groups of 12 rabbits each were established. Groups 1-4 were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 20, 60, 120 and 180 min. Group 5 rabbits received 15 mg/kg bodyweight, i.v., indomethacin and, after 1 h, were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 2 h. Group 6 animals were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 2 h and then received indomethacin. Group 7 rabbits received 100 mg/kg bodyweight, i.v., NG-monomethyl-L-arginine (L-NMMA) and, after 1 h, were ventilated with a mixture of 10% CO2, 21% O2 and 69% N2 for 2 h. Group 8 rabbits were ventilated for 2 h with a mixture of 10% CO2, 21% O2 and 69% N2 and were then administered L-NMMA. Group 9 rabbits received L-NMMA treatment concomitant with ventilation for 2 h with a mixture of 10% CO2, 21% O2 and 69% N2. In all groups, the erythrocyte CA activity was measured, as well as PaCO2 before and after ventilation or treatment. 3. The present study shows that CO2 reduces CA I activity down to complete inhibition and antagonizes the activating effects of indomethacin and L-NMMA on this isozyme. Our data prove that nitric oxide- and prostaglandin-induced CA I inhibition is involved in the vasodilation produced by hypercapnia. These results suggest that, due to subsequent pH changes, CA I is directly implicated in the modulation of vascular processes in the organism.

The mechanism of action of angiotensin II is dependent on direct activation of vascular smooth muscle carbonic anhydrase I.

Our previous studies have shown that angiotensin II increases carbonic anhydrase activity both in vitro and in vivo. In this study we investigated in vitro the effect of angiotensin II on carbonic anhydrase I and II from erythrocytes and on arteriolar vascular smooth muscle carbonic anhydrase I. We also studied in vitro and in vivo the effect of angiotensin II receptor blockers (irbesartan and candesartan) on purified carbonic anhydrase I and II, on vascular smooth muscle carbonic anhydrase I and on arterial blood pressure in humans and in animals. In vitro results showed that angiotensin II is a direct and stronger activator of carbonic anhydrase I than II. Angiotensin II receptor blockers reduced mainly carbonic anhydrase I activity and completely antagonized the activating effect of angiotensin II both on purified and on vascular smooth muscle carbonic anhydrase I. Our in vivo experiments showed that irbesartan and candesartan are powerful inhibitors of carbonic anhydrase I both in erythrocytes (in humans) and in vascular smooth muscles (in animals). In humans, irbesartan and candesartan progressively reduce arterial blood pressure in hypertensive subjects, in parallel with progressive reduction of erythrocyte carbonic anhydrase I activity. We believe that angiotensin II could have a dual mechanism of action: (1) angiotensin interacting with its receptor to form a stimulus-receptor complex; (2) the same stimulus directly acts on the carbonic anhydrase I isozyme (which might be coupled with angiotensin II receptors), ensuring an adequate pH for stimulus-receptor coupling for signal transmission into the cell and hence vasoconstriction.

Erythrocyte superoxide dismutase activity in patients with digest cancer: adjuvant diagnosis test.

Previous studies showed that a common feature of tumor cells is their low of superoxide dismutase (SOD) activity. Our research proved that carcinogenic substances reduce erythrocyte SOD activity, while anticarcinogenic ones increase it and that in vitro SOD assessments revealed a direct mechanism of action. In this paper we determined erythrocyte SOD activity in patients with digestive and extra-digestive cancers. We studied eight groups of patients with five different forms of cancer (histopathologically confirmed) and three groups of volunteers. Group 1 (n = 86), patients with esophageal cancer; Group 2 (n = 94), patients with gastric cancer; Group 3 (n = 79), patients with colorectal cancer; Group 4 (n = 71), patients with hepatic cancer; Group 5 (n = 73), patients with pancreatic cancer; Group 6 (n = 85), patients with with other diseases, but of the same organs; Group 7 (n = 97), healthy volunteers; Group 8 (n = 91), hypertensive patients. We determined the erythrocyte SOD activity using the catecholamine oxidation method. Erythrocyte SOD from the first 5 groups had a mean value of 1.91 +/- 0.78 EU as compared with Group 6 which presented an activity of 6.08 +/- 1.02 (P < 0.001) and Group7 in which the activity was 6.54 +/- 1.23 (P < 0, 001). Erythrocyte SOD activity is significantly reduced (by 2-3 fold) in all groups of cancer patients studied. Being accessible, simple and rapid, erythrocyte SOD activity determination could constitute an adjuvant test in diagnosis of these types of cancer. Taking into account our previous results which prove that carcinogenic substances inhibit SOD activity by a direct mechanism, we suggest that further research is required in this exciting field.

Omeprazole has a dual mechanism of action: it inhibits both H(+)K(+)ATPase and gastric mucosa carbonic anhydrase enzyme in humans (in vitro and in vivo experiments).

In this study our experiments followed in vitro and in vivo the effect of omeprazole on purified and erythrocyte carbonic anhydrase (CA) I and II isozymes, as well as on gastric mucosa CA IV in humans. Our in vitro results show that omeprazole-induced inhibition of purified CA I and CA II and gastric mucosa CA IV is dose- and pH-dependent. In vivo, the i.v. administration of omeprazole in humans in therapeutic doses produced a decrease in erythrocyte CA I and CA II activity, as well as in gastric mucosa CA I, II, and IV. Regarding CA IV, the results lead to the conclusion that omeprazole selectively inhibits gastric mucosa CA IV and does not modify the activity of the same isozyme from the kidney and lung, indicating organ specificity. Our results strongly suggest that omeprazole has a dual mechanism of action: H(+)K(+)ATPase inhibition and gastric mucosa CA inhibition, and that these enzymes may be functionally coupled. This 2-fold mechanism of action could explain the greater effectiveness of substituted benzimidazoles as compared with other therapies.

A new concept regarding the mechanism of action of omeprazole.

OBJECTIVES: In this paper we investigated in humans and in animals the in vitro and in vivo effect of omeprazole upon purified and erythrocyte carbonic anhydrase (CA) I and II isozymes, as well as on gastric mucosa CA IV. METHOD: In vitro, we observed the effect of omeprazole at concentrations between 10(-8)-10(-4) M on purified CA I and CA II, and also on isolated gastric mucosa CA IV, renal and pulmonary CA IV activity, using the dose-response relationship. In vivo, we studied the effect of omeprazole (Losec) on gastric CA I, II and IV, as well as on erythrocyte CA I and CA II, in humans and in animals. RESULTS: In vitro omeprazole inhibits pH-dependent purified CA I and CA II and gastric mucosa CA IV according to dose-response relationship. In vivo, the i.v. administration of omeprazole in rabbits and in humans shows a decrease of erythrocyte CA I and CA II activity as well as of gastric mucosa CA I, II and IV. CONCLUSIONS: Omeprazole in its active form (sulfenamide) selectively inhibits gastric mucosa CA IV and does not modify the activity of the same isozyme from the kidney and lung proving that the enzyme has an organ specificity. Our results lead to the conclusion that omeprazole possesses a dual mechanism of action: both H+K+ATPase and CA inhibition--enzymes that could be in a functional coupling. This dual mechanism of action might explain the higher effectiveness of treatment using substituted benzimidazole inhibitors compared to other therapies.

The inhibitory effect of diuretics on carbonic anhydrases.

A classification of diuretics mainly comprises mercurials; carbonic anhydrase inhibitors, thiazide diuretics, loop diuretics, inhibitors of renal epithelial Na+ channels and antagonists of mineralocorticoid receptors. We studied in this paper the relationship between diuretics and carbonic anhydrase (CA). Our in vitro and in vivo results show that all diuretics inhibit carbonic anhydrase II and renal CA IV. Further, our data show that they also inhibit epithelial cell CA in the renal tubules. The changes in intracellular pH (pHi) induced by these diuretics through CA inhibition would influence: a) the coupling to their receptors affecting information transmission to the epithelial cells of renal tubules as well as diuretic response; b) the decrease of Na+ exchanger (thiazide), of Na+ - K+ - 2Cl- relation (loop diuretics), Na+ channel blocking in distal and collecting tubules (amiloride, triamterene), as well as the antagonism between spironolactone and aldosterone at the mineralocorticoid receptor level, suggest that this competition might also be produced on CA II and on renal CA IV, which, in turn, could be influenced by pH-induced changes, the binding of the diuretic to its membrane receptor as well as the activity of the brush membrane or cytosolic pump. Furosemide and indapamide, diuretics known to have vasodilating effects, induce the fall of blood pressure that parallels the decrease of CA I activity. These results show the involvement of CA in the mechanism of action of the diuretics and in their actions associated with vasodilating effects. pH changes resulting from the action of CA contribute to the action of diuretics. All diuretics inhibit CA isozymes.

Vasodilatory effect of diuretics is dependent on inhibition of vascular smooth muscle carbonic anhydrase by a direct mechanism of action.

Five years ago, our in vitro and in vivo studies demonstrated for the first time that diuretic agents such as furosemide, hydrochlorothiazide, amiloride, triamterene and spironolactone inhibit carbonic anhydrase (CA) I, II and renal CA IV by a direct mechanism of action. In this paper we investigate the relationship between diuretics and CA I in the vasodilatory mechanism. Both in vitro (on purified CA I, erythrocyte CA I and smooth muscle CA I) and in vivo (in human and rabbits) we studied the effect of acetazolamide, hydrochlorothiazide, indapamide, furosemide, amiloride and triamterene on purified CA I, on human erythrocyte CA I, as well as on CA I isolated from vascular smooth muscle. Our results demonstrate that in vitro all diuretics inhibit CA I by a direct mechanism of action. Inhibition reached 100% with acetazolamide, 45% with hydrochlorothiazide, 82% with indapamide, 85% with furosemide, 68% with amiloride and 58% with triamterene. In vivo, similar inhibition of erythrocyte and smooth muscle CA I was obtained, being parallel with a reduction in arterial blood pressure values. Our data show that in addition to their already known mechanisms, diuretics also inhibit CA in vascular smooth muscle. Our results suggest that this mechanism is achieved by means of pH changes induced by CA I inhibition.

Studies on the protective effect of ebrotidine on experimental ulcers induced by non-steroidal anti-inflammatory drugs in healthy volunteers.

Ebrotidine (N-[(E)-[[2-[[[2-[(diaminomethylene)amino]- 4-thiazolyl]methyl]thio]ethyl]amino]methylene]-4-bromo-benzenesulfonamid e, CAS 100981-43-9, FI-3542) is a new H2-receptor antagonist providing a new therapy for the prevention and healing of non-steroidal anti-inflammatory drugs-induced gastroduodenal lesions. Carbonic anhydrase is a zinc enzyme, and its isozyme (carbonic anhydrase II) in parietal cells plays a central role in HCl secretion. The effects of ebrotidine on carbonic anhydrase in human subjects are reported. Eighteen healthy volunteers were distributed in 3 equal subgroups and treated for 10 days as follows: ebrotidine 800 mg/d p.o. (Group A); indometacin 4 mg/kg/d p.o. in 3 divided doses (Group B); ebrotidine 800 mg/d p.o. plus indometacin 4 mg/kg/d p.o. (Group C). Assessment of the enzymatic activity of carbonic anhydrase was based on the colorimetric method of changing pH with the stopped-flow technique. In group A, ebrotidine reduced total gastric mucosal carbonic anhydrase activity by 62%; in group B, indometacin increased carbonic anhydrase activity in gastric mucosa by 138%; in group C, the combined treatment with ebrotidine plus indometacin decreased gastric mucosal carbonic anhydrase activity by 38%. The present study shows that, unlike ranitidine, ebrotidine, a competitive H2-receptor antagonist, is also a non-competitive inhibitor of carbonic anhydrase I and II. By antagonizing the activating effects of indometacin on gastric mucosal carbonic anhydrase, ebrotidine prevents mucosal lesions caused by anti-inflammatory drugs.

Comparative study of the safety and efficacy of ebrotidine versus ranitidine and placebo in the prevention of piroxicam-induced gastroduodenal lesions.

This study assessed the efficacy of ebrotidine (N-[(E)-[[2-[[[2-[(diaminomethylene)amino]-4-thiazolyl] methyl]thio]ethyl]amino]methylene]-4-bromo-benzenesulfonamide, CAS 100981-43-9, FI-3542) versus ranitidine and placebo in preventing gastroduodenal lesions induced by piroxicam. Thirty patients with rheumatic disease, who were divided into 5 groups, received an oral treatment of piroxicam 20 mg once daily for 6 days plus ebrotidine 400 mg/day (Group I); ebrotidine 800 mg/day (Group II); ranitidine 150 mg/day (Group III); ranitidine 300 mg/day (Group IV); or placebo (Group V). Patients were endoscopically examined before and after treatment. Lanza's score was also determined, and laboratory tests were performed. The results of this study showed that the most powerful protective effect against mucosal gastric lesions induced by piroxicam was achieved with 800 mg/day of ebrotidine. Ranitidine at doses of 150 mg/day did not protect gastric mucosa, and the 300 mg/day dose exerted a poor gastroprotective effect.

Isosorbide nitrates, nitroglycerin, and sodium nitroprusside induce vasodilation concomitantly with inhibition of carbonic anhydrase I in erythrocytes.

This study describes the relationship between nitroglycerin, isosorbide dinitrate, sodium nitroprusside, and carbonic anhydrase I, as well as the involvement of this carbonic anhydrase I in vasodilation. Two groups of coronary patients and a group of rabbits underwent treatment with the above-mentioned vasodilating drugs. The activity of red blood cell carbonic anhydrase was monitored and determined by the stopped-flow method. The results show that these drugs inhibit the activity of the isozyme in parallel to their vasodilating effect. The results of this study lead to the hypothesis that through the pH modifications induced by these vasodilators by the inhibition of carbonic anhydrase I, the isozyme may be involved in the regulation of vascular tonus.

Nonsteroidal anti-inflammatory drugs activate carbonic anhydrase by a direct mechanism of action.

Previous studies by this research team proved that vasodilating prostaglandins (PGs) E1, E2 and I2 inhibit carbonic anhydrase (CA) in vitro and in vivo, which suggested involvement of CA in gastric acid secretion inhibition and the increase of gastric mucosa blood flow produced by this group of PGs. Relying on these findings, as well as on our clinical observations, we studied in vitro and in vivo the effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on CA I and CA II. We also followed in vitro the effects on these isozymes of NSAIDs associated to histamine, Ca, PGE2 and acetazolamide. The results show that the NSAIDs used here, which reduce the activity of cyclooxygenase and PG production, activated CA I and CA II in a dose-dependent manner by a mechanism of the noncompetitive type. Histamine and Ca added to NSAIDs amplified the activating effect of the latter on CA II. Association of PGE2 or acetazolamide to NSAIDs reduced NSAID-induced activation of CA I and CA II. Indomethacin abolished the inhibitory effect of acetazolamide on CA I and CA II. Our data imply that between CA and cyclooxygenase there is an inverse relationship, CA activation being accompanied by reduction of cyclooxygenase activity, a reduction achieved by the pH modifications induced by CA activation. In this way, cyclooxygenase, inhibition occurs "via CA," with the pH variations it brings about.

Inhibition of carbonic anhydrase by nitric oxide.

The aim of the present study was to follow the effect of nitric oxide (NO) on carbonic anhydrase in vitro and in vivo. The effect of L-arginine (as source of NO), as well as that of its analogue, nitro-G-monomethyl-L-arginine, an inhibitor of NO synthesis on carbonic anhydrase, were also studied. In vitro results showed that L-arginine activates carbonic anhydrase, while N-G-monomethyl-L-arginine does not modify its activity. In vivo, L-arginine and N-G-monomethyl-L-arginine increased carbonic anhydrase activity by 72% and, 160% respectively. Administration of L-arginine, as a source of NO, and of acetazolamide before administration of N-G-monomethyl-L-arginine abolished the activating effect of the analogue on carbonic anhydrase. These results lead to the conclusion that inhibition of NO synthesis by N-G-monomethyl-L-arginine induces increase of carbonic anhydrase activity. The data also suggest that NO biosynthetized from L-arginine inhibits carbonic anhydrase.

Prostaglandins with vasodilating effects inhibit carbonic anhydrase while vasoconstrictive prostaglandins and leukotriens B4 and C4 increase CA activity.

Previous work carried out by our team in vitro regarding the relationship between prostaglandins (PGs) and carbonic anhydrase (CA) has shown that prostaglandins E1 (PGE1), E2 (PGE2) and I2 (PGI2) inhibit purified bovine red cell CA, as well as human red cell and gastric mucosa CA. This is a completion of our previous research work including kinetic and in vivo studies concerning the relationship between PGs and CA. The relationship between CA and PGE1, E2 and I2, known for their vasodilating and gastric acid secretion (GAS) reducing effects, on the one hand, and between CA and prostaglandin F2 alpha (PGF2 alpha), thromboxans A2 (TXA2), and B4, C4 leukotriens (LTB4, LTC4), known for their vasoconstrictive effects, on the other hand, were studied in vitro. The in vivo studies followed changes induced by vasodilating PGs (misoprostol-analogue of PGE1, nalador-analogue of PGE2, ilomedin-analogue of PGI2) and vasoconstrictive-minprostin analogue of PGF2 alpha on red cell CA, correlated to modifications of arterial blood pressure (BP). Results obtained in vitro show that PGE1, E2, I2 inhibits basal CA activity, while PGF2 alpha, TXA2 and LTB4, LTC4 increase the activity of the enzyme. Kinetic assessments show that the inhibition and the activation mechanism is a direct one of the non-competitive type located on the active site of CA. Data obtained in vivo show that administration of a single therapeutic dose of misoprostol, nalador or ilomedin inhibits basal activity of CA by 35-55% with corresponding decrease of BP values.(ABSTRACT TRUNCATED AT 250 WORDS)